Cytoskeletal mechanics arise not necessarily from the simple sum of individual protein effects, but from the coordinated, emergent actions of multiple components. Traditional single-system assays, such as kinesin-microtubule or myosin-actin interactions, have provided important mechanistic insights but do not capture the collective effects that develop when actin and microtubule filament systems operate together. Actin filaments and microtubules exhibit coordinated coupling mediated through both static and diffusive crosslinkers. The strength and dynamics of this coupling are further governed by environmental factors, including macromolecular crowding and filament density, which modulate both its magnitude and characteristic timescales. To investigate these emergent properties, such as force generation, physical mechanics, and feedback coordination, we developed an in vitro optical tweezers assay that integrates proteins and filaments from both cytoskeletal systems to analyze the force transmission through the system, effective stiffness, relaxation kinetics, and myosin-driven motor activity. Actin-microtubule bundles were assembled using anillin, a dynamic crosslinker that simultaneously binds actin and microtubules and can generate pN-level forces through diffusional expansion. We further modulated crosslinking density by varying anillin concentration and examined how this mechanical coupling is influenced by myosin-driven activity. Together, these perturbations enable quantitative dissection of how crosslinker density and collective motor forces converge to regulate force generation, filament organization, and the emergent mechanics of hybrid actin-microtubule networks.
Chowdhury et al. (Sun,) studied this question.